In order to biochemically define markers of aging in connective tissue several specific hypotheses concerning age-associated changes in collagen crosslinks will be tested. We predict that such changes will include: 1.\a decrease in hydroxylysine containing crosslinks; 2. a decrease in the reducibility of crosslinks, but not necessarily a decrease in the total number of such crosslinks; and 3. an increase in "poly (CB-6)", a collagen-specific peptide. Using newly developed ultra-sensitive HPLC techniques for separation and quantitation of reduced and nonreducible crosslinks, we will analyze skin and lung tissue from rats, humans, and monkeys of various ages. "Poly (CB-6)" will be quantitated by analysis of CNBr peptides prepared from tissue collagen. By examining two different tissues (skin and lung), we will be able to test the hypothesis that the "aging markers" described above occur synchronously in collagens from different tissue sources, even though quantitative relationships may show tissue specificity. By examining three different species, we will be able to compare collagens from subjects of similar temporal and biological age, thus allowing us to attempt to differentiate between changes associated with chronological aging and changes associated with biological aging. Such biochemical markers of tissue aging should have broad relevance to clinical research in gerontology in human subjects.